This project focuses on the mechanisms by which cadherins, integrins and the cytoskeleton cooperate to regulate proliferation in vascular smooth muscle cells (VSMCs). Aberrant dedifferentiation and proliferation of VSMCs plays a major role in vascular stenosis at sites of injury and atherosclerosis. The investigators have shown an aberrant increase in local tissue stiffness at these sites that changes integrin-mediated cell-ECM and cadherin-mediated cell-cell adhesion, both of which appear to be critical for the proliferative sequelae. Understanding how these changes in arterial mechanics regulate proliferation is not only a priority in the development of rational strategies to interrupt progression of vascular disease, but also provides an opportunity to more broadly understand how mechanical forces, cadherins, and integrins cooperate to influence biological functions. We propose that Rho-mediated tension generated in the actin cytoskeleton couples signals from cadherins and integrins in an integrated mechanochemical signaling system that regulates proliferation. In this proposal, the Chen and Assoian labs bring to bear their respective expertise in engineered microenvironments and adhesive regulation of the cell cycle to investigate how tissue stiffening initiates a stimulatory signal for proliferation in an integrated approach that combines in vitro, x vivo, and in vivo model systems. We show that tissue stiffening at sites of vascular injury results in a dramatic increase in N-cadherin expression and that this effect is required for VSMC proliferation and vascular stenosis. Specific Aim 1 will characterize the mechanisms underlying the stimulatory effect of ECM stiffness on N-cadherin and the stimulatory effect of N-cadherin on VSMC cycling. Specific Aim 2 will examine the role of RhoA and cytoskeletal tension in stiffness- and cadherin-induced cycling. To begin to explore the relevance of this novel proliferative pathway in vivo, Specific Aim 3 will examine the interplay between ECM stiffening, FAK and N-cadherin during VSMC cycling in vivo. This project will lead to an integrated molecular understanding of how vascular smooth muscle cells coordinate signals from cadherins, integrins, and cytoskeletal tension into a proliferative response, provide novel approaches to study these complex adhesive and mechanical effects both in vivo and in vitro, and may suggest new therapeutic strategies to interrupt the progression of restenosis and arteriosclerosis.